Rho kinases (ROCK) have been shown to be involved in cellular functions including apoptosis, cell migration, transcriptional activation, fibrosis, cytokinesis, inflammation, and cell proliferation. In neurons ROCK plays a critical role in the inhibition of axonal growth by myelin-associated inhibitory factors such as myelin-associated glycoprotein (MAG). ROCK activity also mediates the collapse of growth cones in developing neurons. Both processes are thought to be mediated by ROCK-induced phosphorylation of substrates such as LIM kinase and myosin light chain phosphatase, resulting in increased contractility of the neuronal actin-myosin system.
Mature neurons preferentially express one of the ROCK isoforms (ROCK 2), which phosphorylates collapsin response mediator protein 2 (CRMP2) that disintegrate growth cones involved in axon branching and elongation in response to stimuli. Inhibiting ROCK 2 prevents expression of CRMP2 that allows growth-cone collapse (Dergham P et al. The Journal of Neuroscience, 22(15):6570-6577, 2002). By suppressing CRMP2, ROCK 2 inhibitors promote neurite expansion, axon elongation, axonal rewiring across lesions within the CNS, and neural regeneration.
Cerebral Ischemia and Stroke
Cerebral ischemia is a deficiency of blood supply to the brain, causing brain damage or an infarction. Cerebral ischemia can be caused by events such as cardiac arrest, systemic hyperperfusion, traumatic brain injury, cerebral thrombosis or hemorrhage (stroke), embolism, near-drowning, arthrosclerosis, birth asphyxia, drug overdose, and hypoxic encephalopathy. Stroke is the third leading cause of death in the U.S. and the resultant brain damage is the leading cause of adult disability as well, because cerebral ischemia can cause brain damage even if blood flow is restored (Kistler, J P et al. Etiology and clinical manifestations of transient ischemic attack. In: UpToDate. Pedley, T A (ed), UpToDate, Wellesley, Mass., 2008). Indeed, such brain damage often occurs after restoration of blood flow to the brain. For example, a component of brain damage from cardiac arrest may not be histologically apparent for approximately 24 to 48 hours after resuscitation from cardiac arrest (U.S. Pat. No. 7,319,090—Methods of Treating Cerebral Ischemia). This delayed brain damage is due to reperfusion disease: i.e., activation of pathological cascades that promote toxic free radical production, release of excitatory amino acids, severe acidosis, and other cellular and molecular changes (Mackay K B et al. Neurodegeneration 5:319-323, 1996). A critical step in this pathway is the rapid accumulation of neutrophils, early-stage inflammation leukocytes that remain present in high concentrations for more than 24 hours after the restoration of blood flow, and it is believed that inhibition of neutrophil infiltration and/or reduction in the amount of circulating neutrophils will lead to improved neurological outcomes (Satoh S et al. Jpn. J. Pharmacol. 80:41-48, 1999).
Treatment for cerebral ischemia is primarily focused on restoring blood flow as soon as a positive diagnosis has been made. The use of anti-platelet and anticoagulant agents is common, including aspirin, heparin, and warfarin (Schievink W I. Curr Opin Cardiol. 15:316, 2000). In extreme cases of large vessel thrombosis, stents or vein grafts may be employed to divert blood flow around the blockage and restore connectivity (Cohen J E et al. Stroke. 34:e254, 2003). Additionally, thrombolytic treatments have been used to promote recanalization if administered within three hours of the ischemic attack, with the risk of additional hemorrhage in the infracted area (The National Institute of Neurological Disorders and Stroke rt-PA Stroke Study Group. N Engl J Med. 333:1581, 1995). In all cases, the cause of the cerebral ischemia is the target of treatment, but the after effects of the inflammation cascade (the major cause of brain infarction from ischemia, second only to the initial hypoxia) are largely ignored—and a rho kinase inhibitor could minimize the neutrophil migration and associated inflammatory response. Furthermore, it is known that rho kinase plays a role in suppressing nerve growth, and thus a rho kinase inhibitor administered after ischemia could not only act as a neuroprotective agent but as a neuroregenerative agent as well (WO 6,072,792 A2—Compounds Which Bind to the Active Site of Protein Kinase Enzymes).
Prophylactic treatments for cerebral ischemia and stroke utilize the above anticoagulants, and are administered to two classes of high-risk patients: first, those with hypertension and a history of cerebrovascular disease (or the condition known as transient ischemic attack), and second, those whose recent (within ten days) ischemic event makes another such event likely.
Spinal Cord Injury
There are nearly 10,000 new cases of spinal cord injury (SCI) each year (The National Spinal Cord Injury Statistical Center. www.spinalcord.uab.edu, accessed May 11, 2007). Traumatic SCI is attributable mainly to auto accidents, falls, acts of violence, or sports in which the vertebrae are fractured, dislocated or compressed. The vertebrae may also be compressed by bleeding, fluid accumulation, and swelling. Non-traumatic SCI is due to arthritis, cancer, inflammation, infection, disc degeneration, or other defects of the spine (Mueller B et al. Nature, 4: 387-397, 2005). Most spinal cord injuries are associated with one or more of the following characteristics: fracture of one or more of the vertebral elements, dislocation of the spinal joint(s), tearing of the supporting ligament structure, and/or disruption/herniation of the intervertebral disc (Sekhon, L H S, et al. Spine, 26:S2, 2001). In its more severe forms, SCI may lead to permanent loss of motor and sensory functions below the site of injury. Quadriplegia is injury at the neck level, below the cervical vertebrae, leading to paralysis of both the arms and legs. Paraplegia is injury at the lower back, below the thoracic and lumbar vertebrae, leading to paralysis of the legs. SCI results from traumatic and non traumatic injuries that damage the nerve fibers in the spinal cord responsible for carrying signals between the brain and the spinal cord, and may affect both the white and gray matter that comprise neurons. Neurons of the central nervous system (CNS) do not readily regenerate or grow after SCI due to three factors: the inherent nature of mature neurons, the extracellular inhibitory environment found in nervous tissue, and the inflammatory environment generated by the trauma (Dergham P et al. The Journal of Neuroscience, 22(15):6570-6577, 2002).
The Rho-ROCK 2 pathway is significantly up-regulated in mammals after spinal cord injury. ROCK 2 inhibitors also function as neuroprotectants by decreasing nervous tissue damage and cavity formation (Mueller B et al. Nature, 4: 387-397, 2005). Inactivation of the Rho-ROCK 2 pathway therefore stimulates and accelerates functional recovery. The activation of the Rho-ROCK 2 pathway also induces apoptosis in neurons and oligonderocytes (Mueller B et al. Nature, 4: 387-397, 2005). Intervention in some or all of the processes with inhibitors of ROCK could reduce the severity of the injury and improve the ultimate outcome of SCI cases.
Neuropathic Pain
Neuropathic pain is chronic pain caused by dysfunction of the peripheral or central nervous system without continuing tissue damage. This includes pain due to neuropathic and idiopathic pain syndromes, and pain associated with neuropathic-related disorders such as cancer, HIV, multiple sclerosis, shingles, spine surgery, diabetic neuropathy, causalgia, brachial plexus avulsion, occipital neuralgia, fibromyalgia, gout, and other forms of neuralgia. Neuropathic pain often involves neural hypersensitivity and can persist without any overt external stimulus. (Goodman & Gilman's “The Pharmacologic Basis of Therapeutics”, 1996, p. 529, McGraw Hill).
The therapeutic objective of most pain therapy is to alleviate the symptoms of pain regardless of the cause. Current pain control therapies include the use of opioids, NSAIDs or ion channel blockers, all of which have safety profiles that are a cause for concern. Individuals with neuropathic disorders and the resulting debilitating neuropathic pain have a decreased quality of life, but agents commonly used to treat other types of pain are usually ineffective, thus there is a need for new agents that are both safe and effective in treating neuropathic pain (Beniczky S et al. J Neural Transm, 112(6):735-49, 2005).
Abnormal activation of the Rho kinase pathway has been demonstrated in various neuropathies (such as brain and spinal cord injuries), resulting in neurite growth inhibition. Rho kinase inhibition results in accelerated regeneration of neurons and enhanced functional recovery after neural injury, thereby potentially preventing neurodegeneration and stimulating neuroregeneration in various neurological disorders (Mueller B K et al. Nat Rev Drug Discovery, 4(5):387-98, 2005; Madura T et al. Plast Reconstr Surg, 119(2):526-35, 2007). It has been demonstrated that Rho kinase is involved in inflammatory pain and the maintenance of neuropathic pain through phosphorylation of myristoylated alanine-rich C-kinase substrate (MARCKS), which may be involved in cytoskeletal restructuring, such as synaptic trafficking and neurotransmitter release. A phosphorylation-site specific antibody against Ser159-phospho-MARCKS (pS159-Mar-Ab) revealed that MARCKS is phosphorylated at Ser159 by Rho kinase and that its phosphorylation is inhibited by a Rho kinase specific inhibitor (Tatsumi S et al. Neuroscience, 131(2):491-8, 2005; Büyükafar K et al. Eur J Pharmacol, 541(1-2):49-52, 2006)
Alzheimer's Disease
Alzheimer's Disease (AD) is a dementing disorder characterized by progressive impairments in memory and cognition. It typically occurs in later life, and is associated with a multiplicity of structural, chemical and functional abnormalities involving brain regions concerned with cognition and memory. Alzheimer's disease is characterized by neurofibrillary tangles (NFTs) and by extracellular amyloid aggregates. The disease typically begins in patients between 60 to 80 years old and progresses to dementia within 5 years and death in approximately 10 years. Current first-line therapies for Alzheimer's disease are cholinesterase inhibitors which enhance the half-life of the acetylcholine in cholinergic synapses involved in learning and memory. NMDA antagonists have recently been approved and are thought to work by decreasing NMDA associated excitotocity. (Alexander, M et al. Treatment of Dementia. In: UpToDate, Rose, B D (Ed), UpToDate, Wellesley, Mass., 2008). Despite the availability of these agents, AD continues to be a debilitating disease and new treatment options are needed. It is therefore clear that there remains today a long standing need for a treatment of AD before the disease has manifested far enough to produce psychological changes, thereby allowing earlier and more effective therapeutic intervention. Furthermore, these treatments do not address the underlying cause of the disease. Alzheimer's has been linked to the toxic 42-amino acid long amyloid −β (Aβ) peptides, as the primary cause of amyloid aggregates, and one possible pathway to lowering the levels of Aβ42 is via certain rho kinase (ROCK) inhibitors, not Formula I or II of the present invention (Mueller B K et al. Nat Rev Drug Discovery, 4(5): 387-98, 2005).
Multiple Sclerosis
Demyelinating diseases are those in which the main pathogenic process causes the destruction of the myelin sheath, which is necessary for the integrity of central nervous system cells. Among demyelinating diseases, multiple sclerosis (MS) is the most frequent disease due to alteration of the myelin in the central nervous system and, with the exception of trauma, it is the most frequent cause of neurological impairment in young adults. It affects 1.5 million people worldwide, and its symptoms generally occur in young adults, therefore its consequences at a personal and socioeconomic level are very severe. (Noseworthy et al., New Engl. J. Med., 343:938-952, 2000) Susceptibility to MS is due to unknown genetic and environmental factors.
There is a consensus among MS researchers according to which the disease has two stages, an initial inflammatory phase of an autoimmune nature, followed by a secondary progressive neurodegenerative phase. In the first phase, activated T cells cross the hematoencephalic barrier, and once inside the central nervous system, they release proinflammatory cytokines triggering an immunological cascade ending in the destruction of the myelin and death of the oligodendrocytes. Knowledge of the autoimmune process with certain detail has served to develop agents of an immuno-modulating nature, the therapeutic efficacy of which is very modest. Until now, different targets for intervention during the inflammatory phase of MS (Zamvil et al., Neuron 38:685-688, 2003) have been disclosed. Among them are those which are focused on reducing inflammation of the nervous system initiated by the activation of the myelin-specific T cells, promoting autoimmunity particularly against components of the myelin, entering the central nervous tissue and releasing in it pro-inflammatory cytokines such as interferon-γ and tumor-α necrosis factor. The immuno-modulator interferon-1β, approved for the treatment of remitting-recurrent MS, also prevents cellular interactions leading to the penetration of activated T cells through the vascular endothelium. Other treatments in clinical trial phase are focused on neutralizing the activity of proinflammatory cytokines and/or to enhance anti-inflammatory ones. However, no medication has been generated which delays or stops the progression of the neurodegenerative phase of the disease which takes a course with progressive neurological degeneration, and which is characterized by the occurrence of severe demyelinating lesions in the white substance with massive oligodendrocyte loss, atrophy and severe axonal damage.
Recent work also implicates p75 in the regulation of axon elongation. Nerve growth factor (NGF) stimulates neurite outgrowth from embryonic rat hippocampal neurons and chick ciliary neurons, which express only p75 for NGF receptors (Yamashita et al., Neuron 24:585-593, 1999). These effects can be accounted for the modulation of Rho activity by p75. Rho is a small GTPase that regulates the state of actin polymerization. In its active GTP-bound form, Rho rigidifies the actin cytoskeleton, thereby inhibiting axonal elongation and mediating growth cone collapse ((Davies, A M, Curr. Biol., 10:R198-200, 2000) & (Schmidt et al., Genes Dev., 16:1587-1609, 2002)). Neurotrophin binding to p75 inactivates RhoA in HN10e cells as well as cerebellar neurons, whereas the over-expression of RhoA in the transfected 293 cells results in the activation of RhoA, suggesting that p75 elicits bi-directional signals. Subsequent study shows that myelin-associated glycoprotein (MAG), a glycoprotein derived from myelin, activates RhoA by a p75-dependent mechanism, thus inhibiting neurite outgrowth from postnatal sensory neurons and cerebellar neurons (Yamashita et al., J. Cell Biol. 157:565-570, 2002). Furthermore, Nogo and oligodendrocyte myelin glycoprotein (OMgp), the other myelin-derived inhibitors of the neurite outgrowth, act on neurons via p75 (Wang et al., Nature 420:74-78, 2002). p75 in complex with the Nogo receptor is suggested to form a receptor for all the myelin-derived inhibitors found so far (Wang et al., Nat Neurosci. 5:1302-1308, 2002). However, the precise mechanism of the regulation of Rho activity by p75 remained to be elucidated.
MS is not curable, and as such, treatments developed to date have focused on slowing the progression of the disease or moderating its symptoms. One treatment includes medicating the patients with either interferon beta-1b or an alternative, glatiramer, both of which will block the immune system's attack on myelin. The following are medications that treat the symptoms of MS: corticosteroids that will reduce inflammation of the nerve tissue, muscle relaxants, and amantadine and modafinil that will reduce fatigue (The Mayo Clinic-MS Treatments [online] 2008 [cited June 18] www.mayoclinic.org/multiple-sclerosis/treatment.html).